Methods and Results: Vascular senescence, inflammation and neointimal formation were measured in iliac arteries of young and old rats at 3, 7, 14 and 30 days after balloon angioplasty. Senescent cells were identified using (SA)-β-gal activity and immunohistochemistry (IHC) for p27. Cell lineage was determined by immunofluorescence confocal microscopy. Inflammatory cells (macrophages, CD68+) appeared in the adventitia 24 h after injury in both groups, disappeared in the young after 3 days, while remaining in remodeled arteries of old animals until 30 days post-surgery. Senescent cells appeared later in the vasculature peaking in number at two weeks after surgery. Arteries of old animals had a higher density of senescent cells than those of young rats (41% ± 14 vs. 5.4% ± 4.17, p=0.0034, n=5). Old animals also developed thicker neointimas at 30 days after injury than their younger counter parts (I/M: 0.8 ± 0.2 vs. 0.54 ± 0.15, p=0.008). Senescent cells (SA-β-gal+) found in injured arteries included vascular smooth muscle cells (VSMC, SMA+ SA-β-gal+), and fibroblasts. Few macrophages (CD68+), lymphocytes (CD4+ or CD8+) and dendritic cells (OX62+) cells were found to be senescent. In addition to this, senescent cells after injury accumulated more DNA adducts and genomic 8-oxo-G, indicating that these cells were under intense oxidative stress. To demonstrate that senescence worsens intimal hyperplasia after injury, matrigel-embedded senescent and non-senescent VSMC were perivascularly seeded around injured vessels. Neointimal formation was dramatically increased in arteries treated with senescent cells when compare to those receiving normal cells (I/M: 0.41 ± 0.105 vs. 0.26 ± 0.04 p=0.014).

Conclusions: These results demonstrate that vascular senescence is not only a consequence of post-injury proliferation and oxidative stress but is also a worsening factor for neointimal development.

This research has received full or partial funding support from the American Heart Association, AHA Greater Southeast Affiliate (Alabama, Florida, Georgia, Louisiana, Mississippi, Puerto Rico & Tennessee).